Electrochemical coagulation assay and device

ABSTRACT

Methods and devices for electrochemically detecting a change in the viscosity of a fluid are provided. In the subject methods, a fluid sample is introduced into an electrochemical cell having oppositely spaced apart working and reference electrodes. An electric potential is applied to the cell to first achieve a steady state cell current. A decrease in the steady state cell current is then detected and related to a change in viscosity of the sample. In many embodiments, the sample is blood and the change in viscosity is related to the onset of coagulation in the blood sample, and often the PT of the blood sample. Also provided are test strips, kits thereof and meters for use in practicing the subject methods.

FIELD OF THE INVENTION

[0001] The field of this invention is coagulation, and particularlycoagulation testing.

BACKGROUND

[0002] Coagulation is defined as a transformation of a liquid or solinto a soft, semi-solid or solid mass. Blood naturally coagulates toform a barrier when trauma or pathologic conditions cause vessel damage.There are two well-recognized coagulation pathways: the extrinsic orthromboplastin-controlled and the intrinsic orprothrombin/fibrinogen-controlled coagulation pathway. Both theextrinsic and intrinsic pathways result in the production of thrombin, aproteolytic enzyme which catalyzes the conversion of fibrinogen tofibrin.

[0003] Coagulation tests which measure a blood sample's ability to forma clot or coagulate have been developed and used to measure theProthrombin Time (PT) of a blood sample. Such tests are commonlyreferred to as PT tests. PT tests find use in a number of differentapplications. For example, PT tests find use in monitoring patientsundergoing anticoagulant therapy. Other situations where PT tests finduse include tests to determine: acquired platelet function defect;congenital platelet function defects; congenital protein C or Sdeficiency; deep intracerebral hemorrhage; DIC (Disseminatedintravascular coagulation); factor II deficiency; factor V deficiency;factor VII deficiency; factor X deficiency; hemolytic-uremic syndrome(HUS); hemophilia A; hemophilia B; hemorrhagic stroke; hepaticencephalopathy; hepatorenal syndrome; hypertensive intracerebralhemorrhage; idiopathic thrombocytopenic purpura (ITP); intracerebralhemorrhage; lobar intracerebral hemorrhage; placenta abruption;transient ischemic attack (TIA); Wilson's disease; and the like. Assuch, PT tests find use in a variety of different applications.

[0004] A number of different PT determination tests and devices havebeen developed. Such devices and test protocols include both opticalbased devices, such as those described in U.S. Pat. No. 6,084,660; to R.Shartle; and electrochemical based devices, such as those described inU.S. Pat. Nos. 6,046,051; 6,060,323 and 6,066,504; all to A. Jina. Inthis latter group of patents a device is disclosed which is suitable forelectrochemical determination of a change of fluid viscosity in asample, where the device is characterized by the presence ofside-by-side electrodes. This configuration requires the use ofrelatively large volumes of sample and a measurement protocol thatimplements a time dependent deconvolution of the background response;i.e., signal is measured over time and is then distinguished overbackground. Thus, the protocols employed with Jina's devices are morecomplicated and perhaps less robust than the protocols used in thepresent invention described below.

[0005] While a number of different PT determination tests and deviceshave been developed, there continues to be a need for additionalprotocols and devices. Of particular interest would be the developmentof PT system that provided for rapid and accurate PT determinations withsmall sample volumes using inexpensive device components, such asdisposable reagent strips. Of even greater interest would be thedevelopment of an electrochemical device and protocol which exhibits theabove desirable parameters, is suitable for use with small samplevolumes and can provide a simple-to-interpret signal that converges to asteady-state value.

[0006] Relevant Literature

[0007] United States Patent of interest include: U.S. Pat. Nos.6,084,660; 6,066,504; 6,060,323; 6,046,051; 5,942,102; 5,916,522;5,628,961; 5,554,531; and 5,300,779. Also of interest are WO 97/18465;WO 95/06868; EP 974840 and GB 1 299 363.

SUMMARY OF THE INVENTION

[0008] Methods and devices for electrochemically detecting a change inthe viscosity of a fluid are provided. In the subject methods, a fluidsample is introduced into an electrochemical cell having oppositelyspaced apart working and reference electrodes. An electric potential isapplied to the cell to first achieve a steady state cell current. Adecrease in the steady state cell current is then detected and relatedto a change in viscosity of the sample. In many embodiments, the sampleis blood and the change in viscosity is related to the onset ofcoagulation in the blood sample, and often the PT of the blood sample.Also provided are test strips, kits thereof and meters for use inpracticing the subject methods.

BRIEF DESCRIPTION OF THE FIGURES

[0009]FIG. 1 provides an exploded view of a reagent test strip accordingto the subject invention.

[0010]FIG. 2 shows the time-current plot of a typical data set whereblood is introduced into a strip and the current is monitored with time.

DESCRIPTION OF THE SPECIFIC EMBODIMENTS

[0011] Methods and devices for electrochemically detecting a change inthe viscosity of a fluid are provided. In the subject methods, a fluidsample is introduced into an electrochemical cell having oppositelyspaced apart working and reference electrodes. An electric potential isapplied to the cell to first achieve a steady state cell current. Adecrease in the steady state cell current is then detected and relatedto a change in viscosity of the sample. In many embodiments, the sampleis blood and the change in viscosity is related to the onset ofcoagulation in the blood sample, and often the PT of the blood sample.Also provided are test strips, kits thereof and meters for use inpracticing the subject methods.

[0012] Before the subject invention is described further, it is to beunderstood that the invention is not limited to the particularembodiments of the invention described below, as variations of theparticular embodiments may be made and still fall within the scope ofthe appended claims. It is also to be understood that the terminologyemployed is for the purpose of describing particular embodiments, and isnot intended to be limiting. Instead, the scope of the present inventionwill be established by the appended claims.

[0013] In this specification and the appended claims, singularreferences include the plural, unless the context clearly dictatesotherwise. Unless defined otherwise, all technical and scientific termsused herein have the same meaning as commonly understood to one ofordinary skill in the art to which this invention belongs.

[0014] Methods

[0015] As summarized above, the subject invention provides a method fordetermining a change in viscosity of a fluid sample. Often the subjectmethods provide a means for determining or detecting an increase in theviscosity of a fluid sample. The subject methods are sufficientlysensitive to detect an increase in viscosity that is less than about 1cps, and often less than about 0.5 cps in magnitude. As such, thesubject methods are sensitive methods for detecting a change inviscosity of a fluid sample.

[0016] Another feature of the subject methods is that they areelectrochemical methods for determining a change, and often an increase,in the viscosity of a fluid sample. By electrochemical methods is meantthat the subject methods employ a working and a reference electrode.Specifically, the subject methods employ a current produced between aworking and reference electrode and changes therein to determine achange in viscosity of the fluid sample, as described in greater detailbelow.

[0017] The first step in the subject methods is to introduce a quantityof the fluid to be assayed, i.e., a fluid sample, into anelectrochemical cell that includes oppositely spaced apart working andreference electrodes. The nature of the fluid may vary, so long as thefluid is a conductor, e.g., an electrolyte. In many embodiments, thefluid is an aqueous fluid, where of particular interest arephysiological samples. Where the fluid is a physiological sample, inmany embodiments the fluid is whole blood, or a derivative thereof fromwhich the coagulation/clotting time, and therefore PT time, can bederived.

[0018] The amount of fluid, e.g., blood, that is introduced into theelectrochemical cell varies, but is generally a small volume. As such,the volume of fluid introduced into the electrochemical cell typicallyranges from about 0.1 to 10 μL, usually from about 0.2 to 5.0 μL, andmore usually from about 0.3 to 1.6 μL. The sample is introduced into theelectrochemical cell using any convenient protocol, where the sample maybe injected into the electrochemical cell, allowed to wick into theelectrochemical cell, and the like, as may be convenient and dependingon the nature of the device/system in which the subject method ispracticed.

[0019] While the subject methods may be used, in principle, with anytype of electrochemical cell having oppositely spaced apart working andreference electrodes, in many embodiments the subject methods employ anelectrochemical test strip. The electrochemical test strips employed inthese embodiments of the subject invention are made up of two opposingmetal electrodes separated by a thin spacer layer, where thesecomponents define a reaction area or zone that makes up theelectrochemical cell.

[0020] In certain embodiments of these electrochemical test strips, theworking and reference electrodes are generally configured in the form ofelongated rectangular strips. Typically, the length of the electrodesranges from about 1.9 to 4.5 cm, usually from about 2.0 to 2.8 cm. Thewidth of the electrodes ranges from about 0.07 to 0.76 cm, usually fromabout 0.24 to 0.60 cm. The working and reference electrodes typicallyhave a thickness ranging from about 10 to 100 nm and usually from about10 to 20 nm. FIG. 1 provides an exploded view of an electrochemical teststrip according to the subject invention.

[0021] The working and reference electrodes are further characterized inthat at least the surface of the electrodes that faces the reaction areaof the electrochemical cell in the strip is a metal, where metals ofinterest include palladium, gold, platinum, silver, iridium, carbon(conductive carbon ink), doped tin oxide, stainless steel and the like.In many embodiments, the metal is gold or palladium. While in principlethe entire electrode may be made of the metal, each of the electrodes isgenerally made up of an inert support material on the surface of whichis present a thin layer of the metal component of the electrode. Inthese more common embodiments, the thickness of the inert backingmaterial typically ranges from about 25 to 500, usually 50 to 400 μm,e.g., from about 127 to 178 μm, while the thickness of the metal layertypically ranges from about 10 to 100 nm and usually from about 10 to 40nm, e.g. a sputtered metal layer. Any convenient inert backing materialmay be employed in the subject electrodes, where typically the materialis a rigid material that is capable of providing structural support tothe electrode and, in turn, the electrochemical test strip as a whole.Suitable materials that may be employed as the backing substrate includeplastics, e.g. PET, PETG, polyimide, polycarbonate, polystyrene,silicon, ceramic, glass, and the like.

[0022] A feature of the electrochemical test strips used in theseembodiments of the subject methods is that the working and referenceelectrodes as described above face each other and are separated by onlya short distance, such that the distance between the working andreference electrodes in the reaction zone or area of the electrochemicaltest strip is extremely small. This minimal spacing of the working andreference electrodes in the subject test strips is a result of thepresence of a thin spacer layer positioned or sandwiched between theworking and reference electrodes. The thickness of this spacer layer mayrange from 50 to 750 μm and is often less than or equal to 500 μm, andusually ranges from about 100 to 175 μm, e.g., 102 to 153 μm. The spacerlayer is cut so as to provide a reaction zone or area with at least aninlet port into the reaction zone, and generally an outlet port out ofthe reaction zone as well. The spacer layer may have a circular reactionarea cut with side inlet and outlet vents or ports, or otherconfigurations, e.g. square, triangular, rectangular, irregular shapedreaction areas, etc. The spacer layer may be fabricated from anyconvenient material, where representative suitable materials includePET, PETG, polyimide, polycarbonate, and the like, where the surfaces ofthe spacer layer may be treated so as to be adhesive with respect totheir respective electrodes and thereby maintain the structure of theelectrochemical test strip. Of particular interest is the use of adie-cut double-sided adhesive strip as the spacer layer.

[0023] The electrochemical test strips used in these embodiments of thesubject invention include a reaction zone or area that is defined by theworking electrode, the reference electrode and the spacer layer, wherethese elements are described above. Specifically, the working andreference electrodes define the top and bottom of the reaction area,while the spacer layer defines the walls of the reaction area. Thevolume of the reaction area typically ranges from about 0.1 to 10 μL,usually from about 0.2 to 5.0 μL, and more usually from about 0.3 to 1.6μL. As mentioned above, the reaction area generally includes at least aninlet port, and in many embodiments also includes an outlet port. Thecross-sectional area of the inlet and outlet ports may vary as long asit is sufficiently large to provide an effective entrance or exit offluid from the reaction area, but generally ranges from about 9×10⁻⁴ to5×10⁻³ cm², usually from about 1.3×10⁻³ to 2.5×10⁻³ cm².

[0024] In many embodiments, a reagent system is present in the reactionarea, where the reagent system interacts with components in the fluidsample during the assay. For example, in embodiments where the subjectmethods are used to detect a coagulation event, e.g., to measure PT of asample, the reaction area or zone includes a reagent system that atleast includes a redox couple, and often also includes a coagulationcatalyzing agent.

[0025] The redox couple of the reagent composition, when present, ismade up of one or more redox couple agents. A variety of different redoxcouple agents are known in the art and include: ferricyanide, phenazineethosulphate, phenazine methosulfate, pheylenediamine,1-methoxy-phenazine methosulfate, 2,6-dimethyl-1,4-benzoquinone,2,5-dichloro-1,4-benzoquinone, ferrocene derivatives, osmium bipyridylcomplexes, ruthenium complexes, and the like. In many embodiments, redoxcouples of particular interest are ferricyanide, and the like.

[0026] In many embodiments, the reagent composition also includes acoagulation catalyzing agent. By coagulation catalyzing agent is meantone or more components or reactants that participate or interact withcomponents present in the fluid sample, e.g., whole blood, to initiatethe clotting process in the blood sample. For PT assays, the coagulationcatalyzing agent generally comprises thromboplastin, whichthromboplastin may be purified from a naturally occurring source, e.g.,an aqueous extract of acetone dried brain tissue, or syntheticrecombinant thromboplastin (r-DNA thromboplastin), which generallyincludes purified recombinant tissue factor protein and a purifiedartificial lipid component. A representative coagulation catalyzingagent is thromboplastin-XS with calcium sold under the trade nameINNOVIN® by Dade International, Miami Fla.

[0027] Other reagents that may be present in the reaction area includebuffering agents, e.g. citraconate, citrate, malic, maleic, phosphate,“Good” buffers and the like. Yet other agents that may be presentinclude: divalent cations such as calcium chloride, and magnesiumchloride; surfactants such as Triton, Macol, Tetronic, Silwet, Zonyl,and Pluronic; stabilizing agents such as albumin, sucrose, trehalose,mannitol, and lactose.

[0028] The reagent system, when present, is generally present in dryform. The amounts of the various components may vary, where the amountof the oxidized redox couple component typically ranges from about 5 to1000 mM, usually from about 90 to 900 mM; the reduced redox couplecomponent typically ranges from about 1 to 20 mM, usually from about 5to 15 mM; the amount of buffer typically ranges from about 0 to 300 mM,usually from about 50 to 100 mM; and the amount of coagulationcatalyzing agent component typically ranges from about 0.005 to 50mg/cm², usually from about 0.05 to 5 mg/cm². The overall mass of dryreagent present in the reaction area or zone in these embodimentsgenerally ranges from about 4 to 700 ng/cm², usually from about 8 to 350ng/cm².

[0029] A representative test strip for use in the subject methods isdepicted in exploded view in FIG. 1.

[0030] Following sample introduction into the electrochemical cell, aconstant electric potential is applied to the cell in a mannersufficient to produce a steady state current between the working andreference electrodes of the cell. More specifically, a constant electricpotential is applied between the working and the reference electrodes ina manner that produces a steady state current between the twoelectrodes. The magnitude of the applied electric potential generallyranges from about 0 to −0.6 V, usually from about −0.2 to −0.4 V. Inmany embodiments where the electrochemical cell includes a redox couple,as described above, application of the constant electrical potential asdescribed above results in the production of a steady state currentdescribed by the following formula:

i_(ss)=n2FADCo/L;

[0031] where:

[0032] n is equal to the number of electrons transferred;

[0033] F is Faraday's constant, i.e., 9.6485×10⁴C/mol;

[0034] A is the area of the working electrode;

[0035] D is the diffusion coefficient of the cell, where thiscoefficient may be determined from Fick's first law, i.e.J(x,t)=−D^(dCo(x,t))/dx where j is flux, x is the position from theelectrode, and t is time;

[0036] Co is the redox couple concentration, e.g., the ferrocyanideconcentration; and

[0037] L is distance between the electrodes, e.g., the spacer thickness.

[0038] The overall time period required to obtain the requisite steadystate current, as described above, is relatively short in certainembodiments. In such embodiments, the total amount of time required toobtain the steady state current, i.e., the period from sample entry tothe cell to establishment of the steady state current, is less thanabout 15 seconds, usually less than about 10 seconds; and often rangesfrom about 4 to 15 seconds.

[0039]FIG. 2 shows the time-current plot of a typical data set whereblood is introduced into a strip and the current is monitored with time.

[0040] The next step in the subject methods is to detect a change in thesteady state current and relate this change to a change in viscosity ofthe sample. In many embodiments, the change that is detected is adecrease in the steady state current. The magnitude of the decrease inthe steady state current that is detected in this step is at least about2%, and usually at least about 10%, where the magnitude of the decreasein many embodiments ranges from about 2 to 90%. In other embodiments, ofinterest is the rate of change between two steady state values, onebefore and one after the coagulation event, and the relation of thischange in rate to the presence of the coagulation event.

[0041] The detection of the above described decrease in steady statecurrent is then related to an increase in viscosity of the fluid samplein the electrochemical cell. Relatively small increases in viscosityresult in a detectable decrease in the steady state current and thus canbe detected by the subject methods, where the magnitude of the increasein viscosity may be as small as 0.5 cps or smaller in certainembodiments.

[0042] In many embodiments where the sample present in theelectrochemical cell is whole blood and the reagent composition includesa coagulation catalyzing agent, the increase in viscosity is thenrelated to the onset of coagulation in the blood sample, i.e., theoccurrence of a coagulation event or blood clotting in the blood sample.

[0043] In certain embodiments, the increase in viscosity and concomitantdetection of the onset of coagulation in the blood sample being assayedis employed to determine the PT of the blood sample. In theseembodiments, the period extending from the initial sample introductioninto the reaction area or zone and/or the establishment of a steadystate current and increase in viscosity/onset of coagulation isdetermined and the PT of the blood sample is derived from this timeperiod. The time at which sample enters the electrochemical cell may bedetected using any convenient protocol, where particular protocolsemployed may depend, at least in part, on the nature of the meter deviceemployed with the electrochemical cell. In certain embodiments, the timethat sample is introduced directly into the reaction cell can bemanually recorded. Alternatively, the meter may automatically detectsample introduction into the electrochemical cell, e.g., by detecting aninitial decrease in the voltage required to achieve a constant currentbetween the working and reference electrodes of the cell. (See U.S.application Ser. No. 9/333,793, filed Jun. 15, 1999, incorporated hereinby reference.) Other protocols for sample detection in the cell may alsobe employed.

[0044] The above computational steps of the subject method, e.g.,relation of the time period from sample introduction to onset ofcoagulation to the PT of the blood sample, may be accomplished manuallyor through the use of an automated computing means, where in manyembodiments the use of an automated computing means, such as isdescribed in connection with the subject devices discussed below, is ofinterest.

[0045] The above described protocol may be carried out at roomtemperature or at an elevated temperature. Typically, the above protocolis carried out at a temperature ranging from about 20 to 40° C., e.g.,about 37° C.

[0046] The above described methods find use in any application where thedetermination of a viscosity change in a fluid sample is desirable. Assuch, the subject methods suited for use in the determination of PT of ablood sample, and as such find use in any application where thedetermination of PT is desired, e.g., those applications described inthe Background Section, supra.

[0047] Devices

[0048] Also provided by the subject invention are meters for use inpracticing the subject invention. The subject meters are typicallymeters for measuring a change in viscosity of fluid sample, and aremeters for measuring the PT of a blood sample in many embodiments. Thesubject meters typically include: (a) a means for applying an electricpotential to an electrochemical cell into which the sample has beenintroduced; (b) a means for measuring cell current in the cell,including a steady state current in the cell; (c) a means for detectinga change in the steady state current in the cell, e.g., a decrease inthe steady state current of the cell; and (d) a means for relating thechange in steady state current to a change in viscosity of the cell,e.g., a means for relating a decrease in steady state current in thecell to an increase in viscosity of fluid in the cell.

[0049] The means for applying an electric potential to theelectrochemical cell, means for measuring a steady state current in thecell and means for detecting a change in the steady state current in thecell may be any convenient means, where representative means aredescribed in WO 97/18465 and U.S. Pat. No. 5,942,102; the disclosures ofwhich are herein incorporated by reference. See also U.S. Pat. Nos.6,066,504; 6,060,323; 6,046,051; the disclosures of which are hereinincorporated by reference. The means for relating the change in steadystate current to a change in viscosity is typically a computing meanspresent in the meter which is capable of relating the measured change insteady state current to a change in viscosity of the fluid sample. Inmany embodiments, this means is further a means for relating the changein current/viscosity to the onset of coagulation, and is often a meansfor determining the PT of a blood sample. See e.g., U.S. Pat. No.6,066,504; the disclosure of which is herein incorporated by reference.

[0050] Kits

[0051] Also provided are kits for use in practicing the subject methods.The kits of the subject invention at least include an electrochemicalreagent test strip, as described above. The subject kits may furtherinclude a means for obtaining a physiological sample. For example, wherethe physiological sample is blood, the subject kits may further includea means for obtaining a blood sample, such as a lance for sticking afinger, a lance actuation means, and the like. In addition, the subjectkits may include a calibration means for calibrating the instrument,e.g., a control solution or standard, e.g., a coagulation controlsolution that has a known PT time. In certain embodiments, the kits alsoinclude an automated instrument, as described above, for detecting theamount of product produced on the strip following sample application andrelating the detected product to the amount of analyte in the sample.Finally, the kits include instructions for using the subject kitcomponents in the determination of an analyte concentration in aphysiological sample. These instructions may be present on one or moreof the packaging, a label insert, containers present in the kits, andthe like.

[0052] The following examples are offered by way of illustration and notby way of limitation.

Experimental

[0053] I. Electrochemical Test Strip Preparation

[0054] An electrochemical test strip consisting of two metallizedelectrodes oriented in a sandwich configuration was prepared as follows.The top layer of the test strip was a gold sputtered Mylar strip. Themiddle layer was a double-sided adhesive with a punched hole thatdefined the reaction zone or area. The punched hole was a circle withtwo juxtaposed rectangular inlet and outlet channels. The bottom layerof the test strip was sputtered palladium on Mylar. A reagent ofcitraconate buffer, ferricyanide, ferrocyanide and relipidatedrecombinant tissue factor was ink jetted on the palladium sputteredsurface. The amount of reagent ink jetted onto the palladium sputteredsurface was 597 ng/cm². As such, the amount of citraconate buffer was120 ng/cm² ferricyanide was 460 ng/cm², the amount of ferrocyanide was 8ng/cm² and the amount of recombinant tissue factor was 9 ng/cm². Anexploded view of the test strip is shown in FIG. 1.

[0055] II. Detection of PT

[0056] The above described strip is employed to determine the PT of ablood sample as follows. A 1.5 μl blood sample is introduced into thereaction area or zone of the test strip and the sample introduction timeis recorded. A constant potential of −0.3 V is applied between theworking and reference electrodes, and the resultant current between thetwo electrodes is monitored. The appearance of a steady state current isfirst detected, followed by a decrease in the steady state current. Thetime period from the initial sample introduction to the decrease insteady state current is determined and then related to the PT of theblood sample. FIG. 2 shows the time-current plot of the data set whereblood is introduced into a strip and the current is monitored with time.

[0057] The above results and discussion demonstrate that subjectinvention provides a simple and powerful tool to determine the PT of ablood sample. Advantages of the subject methods over non-electrochemicalbased coagulation detection methods include use of low cost materialsand the opportunity to use wide variety of controls, including plasmabased controls. Additional advantages of the subject invention includethe ability to employ small sample volumes and the fact that theelectrochemical measurements made by the subject methods provide asimple-to-interpret signal that converges to a steady-state value. Yetanother advantage is the ability to use low cost electrochemical basedmeters, which provide for significant cost savings. As such, the subjectinvention represents a significant contribution to the art.

[0058] All publications and patents cited in this specification areherein incorporated by reference as if each individual publication orpatent were specifically and individually indicated to be incorporatedby reference. The citation of any publication is for its disclosureprior to the filing date and should not be construed as an admissionthat the present invention is not entitled to antedate such publicationby virtue of prior invention.

[0059] Although the foregoing invention has been described in somedetail by way of illustration and example for purposes of clarity ofunderstanding, it is readily apparent to those of ordinary skill in theart in light of the teachings of this invention that certain changes andmodifications may be made thereto without departing from the spirit orscope of the appended claims.

What is claimed is:
 1. A method for detecting a change in the viscosityof a fluid sample, said method comprising: (a) introducing a sample intoan electrochemical cell comprising oppositely spaced apart working andreference electrodes; (b) applying an electric potential to saidreaction cell to produce a steady state current between said oppositelyspaced apart electrodes; (c) detecting a change in said steady statecurrent; and (d) relating said change in steady state current to achange in viscosity of said fluid sample.
 2. The method according toclaim 1, wherein said change in steady state current is a decrease insteady state current.
 3. The method according to claim 1, wherein saidchange in viscosity is an increase.
 4. The method according to claim 1,wherein said fluid sample is a physiological sample.
 5. The methodaccording to claim 4, wherein said physiological sample is blood.
 6. Themethod according to claim 5, wherein said method further comprisesrelating said change in viscosity to the prothrombin time (PT) of saidblood.
 7. The method according to claim 1, wherein said electrochemicalcell comprises a redox couple.
 8. A method for detecting the onset ofcoagulation of a blood sample, said method comprising: (a) introducingsaid blood sample into an electrochemical cell comprising: (i)oppositely spaced apart working and reference electrodes; and (ii) areagent mixture comprising a redox couple; (b) applying an electricpotential to said reaction cell to produce a steady state currentbetween said oppositely spaced apart electrodes; (c) detecting a changein said steady state current; and (d) relating said change in steadystate current to the onset of coagulation in said blood sample.
 9. Themethod according to claim 8, wherein said change is a decrease.
 10. Themethod according to claim 8, wherein said reagent comprises acoagulation catalyzing agent.
 11. The method according to claim 10,wherein said coagulation catalyzing agent comprises thromboplastin. 12.The method according to claim 10, wherein said method further comprisesrelating said onset of coagulation to the prothrombin time of said bloodsample.
 13. An electrochemical test strip comprising: an electrochemicalcell comprising: (a) oppositely spaced apart working and referenceelectrodes; and (b) a reagent mixture comprising: (i) a redox couple;and (ii) a coagulation catalyzing agent.
 14. The reagent test stripaccording to claim 13, wherein said oppositely spaced working andreference electrodes are separated by a distance ranging from about 50to 750 μm.
 15. The reagent test strip according to claim 14, whereinsaid coagulation catalyzing agent comprises thromboplastin.
 16. Thereagent test strip according to claim 13, wherein said redox couplecomprises a ferricyanide and ferrocyanide.
 17. The reagent test stripaccording to claim 13, wherein said electrochemical cell has a volumeranging from about 0.1 to 10 μL.
 18. A meter for detecting a change inviscosity of a fluid sample, said meter comprising: (a) means forapplying an electric potential to an electrochemical cell made up ofoppositely spaced apart working and reference electrodes and comprisingsaid fluid sample; (b) means for measuring cell current between saidoppositely space apart working and reference electrodes; (c) means fordetecting a change in said measured cell current; and (d) means forrelating said change in measured cell current to a change in viscosityof said fluid sample.
 19. The meter according to claim 18, wherein saidmeter further comprises a means for relating said change in viscosity tothe prothrombin time of said fluid sample.
 20. A kit for use indetecting a coagulation event in a blood sample, said kit comprising:(a) at least one electrochemical test strip comprising anelectrochemical cell comprising: (i) oppositely spaced apart working andreference electrodes; and (ii) a reagent mixture comprising a redoxcouple and a coagulation catalyzing agent; and (b) at least one of acalibration means and a means for obtaining a sample.